Fig. 2: Ephexin1 enhances the MEK/ERK signaling pathway and regulates K-Ras-downstream target genes.

A Whole-cell lysates from control or Ephexin1-depleted H1299 cells were incubated with human phosphokinase array membranes. Each pair of the most positive kinase dots is numbered with the identification of the corresponding kinase listed. B Levels of indicated protein kinases were quantified after normalized positive control. Error bars indicate mean ± SD of spots. C Western blot analysis to detect MEK/ERK activation in indicated cells treated with EGF for the indicated amounts of time after serum starvation. D The co-expression heat map of Ephexin1 with K-Ras, H-Ras, N-Ras, or Ki-67 in TCGA and GTEx (n = 5151) derived from the UCSC Xena browser. E Heat map showing the differential expressed genes induced by Ephexin1 shRNA in H1299 cells. Red and green indicate high and low mRNA expression levels, respectively. F Scatter plot shows the differently expressed genes (FDR ≤ 0.025 and fold difference ≥ 2) in Ephexin1-depleted H1299 cells compared with control cells. Compared to the control group (shControl), shEphexin1 downregulates 155 genes, whereas upregulates 227 genes expression levels. White circles highlight the position of the indicated K-Ras-downstream target genes. G Gene set enrichment analysis (GSEA) was used to identify K-Ras-downstream target genes differentially expressed between control and Ephexin1-depleted H1299 cells. H Heatmap of Ras-target genes downregulated and upregulated by Ephexin1 knockdown in H1299 cells. I RT-qPCR analysis of indicated genes. Values represent relative expression normalized to β-actin mRNA ± SD. ^**P < 0.01; ^***P < 0.001 compared with control. P values are for a two-tailed Student’s t test.